Normal-appearing Skin Research Articles

Melanoma arising from pre-existing naevus in carriers of a germline CDKN2A pathogenic variant.

In this study we used total body photography (TBP) to determine whether melanoma emerges from a preexisting naevi or normal-appearing skin in individuals with pathogenic CDKN2a variant. In total, 60 melanoma cases occurred in 44 CDKN2a germline carriers. In addition, we observed that 78% of the 60 included melanoma cases developed from preexisting naevi, significantly higher than the 30% reported for sporadic melanoma. This high incidence may be due to the numerous atypical naevi and early bi-allelic loss of CDKN2A. Our findings underscore the importance of routine TBP in these patients for early detection of melanoma. More than previously appreciated, the surgical removal of melanocytic naevi in these high-risk patients may prevent melanoma development.

Intralesional immunotherapy of cutaneous warts using tuberculin purified protein derivative and vitamin D3 – A randomized comparative study

Objectives: Numerous studies have been conducted on the clinical efficacy of immunotherapeutic agents in the treatment of cutaneous warts showing variable results. The present study aimed to compare the clinical efficacy and safety of intralesional tuberculin purified protein derivative (PPD) and Vitamin D3 therapy in recurrent and recalcitrant extra-genital cutaneous warts. Material and Methods: This study was conducted as a prospective, randomized, comparative, and single-blind study. A total of 104 patients were randomly distributed into two treatment groups: Group A (Tuberculin PPD, n = 53) and Group B (Vitamin D3, n = 51). Each patient in Group A received an intralesional injection of 0.1 mL tuberculin PPD (5 Tuberculin Units), while each patient in Group B received an intralesional injection of 0.2 mL Vitamin D3 (Cholecalciferol containing 120,000 IU). The injections were given at the initial visit (week 0) and after every 2 weeks for a maximum of four sessions (weeks 2, 4, and 6). The largest wart was selected for intralesional therapy. The categorization of clinical response was done based on the percent reduction in the size of warts into complete (appearance of normal skin), marked (>50% reduction), moderate (<50% reduction), and inadequate (no change in warts) responses. Adverse effects (if any) were recorded during each patient visit. The final response was evaluated at 6 months follow-up from the last treatment session. Results: Regarding the response of patients to tuberculin PPD therapy, out of a total of 53 patients, 40 (75.5%) showed a complete response. Regarding the response of patients to Vitamin D3 therapy, out of a total of 51 patients, 36 (70.6%) showed a complete response. However, the difference in the response to the treatment between the two groups was statistically insignificant (P = 0.402). Conclusion: Both intralesional tuberculin PPD and Vitamin D3 are effective and safe in the treatment of all recurrent and recalcitrant extra-genital cutaneous warts.

IL-17A/F double producing T cells, unstable Tregs and quiescent TRMs in clinically healed lesions are potential cellular candidates for recurrence of psoriasis

Biological antibodies targeting key cytokines such as IL-17 and IL-23 have revolutionized psoriasis outcome. However, the recurrence remains an urgent challenge to be addressed. Currently, most of the descriptions of skin T-cell characteristics in psoriasis are derived from lesional and non-lesional skin, and their characteristics in resolved lesions (clinically healed lesions) remain vague. In order to further elucidate the cellular mechanism of recurrence, we performed single-cell sequencing and multiplexed immunohistochemical staining of T-cell subsets in autologous resolved lesion (RL), on-site recurrent psoriatic lesion (PL), and adjacent normal-appearing skin (NS) of psoriasis. By comparing with PL and NS tissues, we identified three potential cellular candidates for recurrence in clinically healed lesions: IL-17A/F double producing T cells, unstable Tregs and quiescent TRMs. Our results provide research clues for elucidating the immunological recurrence mechanism of psoriasis, and further work is needed to deepen our findings.

Transcriptomic evaluation of skin tape-strips in children with allergic asthma uncovers epidermal barrier dysfunction and asthma-associated biomarkers abnormalities.

Tape-strips, a minimally invasive method validated for the evaluation of several skin diseases, may help identify asthma-specific biomarkers in the skin of children with allergic asthma. Skin tape-strips were obtained and analyzed with RNA-Seq from children with moderate allergic asthma (MAA) (n = 11, mean age 7.00; SD = 1.67), severe allergic asthma (SAA) (n = 9, mean age 9.11; SD = 2.37), and healthy controls (HCs) (n = 12, mean age 7.36; SD = 2.03). Differentially expressed genes (DEGs) were identified by fold change ≥2 with a false discovery rate <0.05. Transcriptomic biomarkers were analyzed for their accuracy in distinguishing asthma from HCs, their relationships with asthma-related outcomes (exacerbation rate, lung function-FEV1, IOS-R5-20, and lung inflammation-FeNO), and their links to skin (barrier and immune response) and lung (remodeling, metabolism, aging) pathogenetic pathways. RNA-Seq captured 1113 in MAA and 2117 DEGs in SAA. Epidermal transcriptomic biomarkers for terminal differentiation (FLG/filaggrin), cell adhesion (CDH19, JAM2), lipid biosynthesis/metabolism (ACOT2, LOXL2) were significantly downregulated. Gene set variation analysis revealed enrichment of Th1/IFNγ pathways (p < .01). MAA and SAA shared downregulation of G-protein-coupled receptor (OR4A16, TAS1R3), upregulation of TGF-β/ErbB signaling-related (ACVR1B, EGFR, ID1/2), and upregulation of mitochondrial-related (HIGD2A, VDAC3, NDUFB9) genes. Skin transcriptomic biomarkers correlated with the annualized exacerbation rate and with lung function parameters. A two-gene classifier (TSSC4-FAM212B) was able to differentiate asthma from HCs with 100% accuracy. Tape-strips detected epithelial barrier and asthma-associated signatures in normal-appearing skin from children with allergic asthma and may serve as an alternative to invasive approaches for evaluating asthma endotypes.

B-223 Validation of a Non-invasive DNA NGS Assay for Detecting Somatic Mutations in Normal Appearing Sun Exposed Skin

Abstract Background Ultraviolet radiation (UVR) exposure is a major risk factor for developing nonmelanoma skin cancer and somatic mutations caused by UVR exposure can arise even in normal-appearing skin. Non-invasive identification of UVR induced somatic mutations can provide a quantitative evaluation of the extent of UVR damage. This study describes the development and analytical validation of a noninvasive method for identifying low level somatic variants in sun exposed peripheral skin by ultra-deep Next Generation Sequencing, in specimens collected using adhesive tape designed to non-invasively collect skin samples. Methods Five genomic regions with the highest density of known UVR induced somatic mutations across TP53 and NOTCH1 were amplified by 5-plex PCR primers affixed with Illumina NGS adapters. These amplicons were purified, size selected and then indexed using Unique Dual Indexes. The NGS libraries were pooled to 96-plex and bi-directionally sequenced with 2x150 chemistry on the Illumina iSeq100. Data analysis was performed using the Illumina DRAGEN Somatic Variant Calling pipeline. Titrations between 0.01 and 1 of synthetic controls comprising plasmids containing amplimer inserts with 19 of the most well characterized UVR-induced C &amp;gt; T driver mutations spiked into a wildtype insert were sequenced in duplicate. In addition, 3 cell line controls at input concentrations of a 2-fold serial dilution from 13.2 ng–0.4 ng were spiked into a mutation negative cell line at variant allele frequencies between 0.005 and 1 and sequenced in triplicate across three NGS runs. Finally, 178 specimens collected from the central forehead, nose and cheeks of human donors were sequenced and their cumulative mutation frequency analyzed by linear regression against the age of the participants. Results All samples were sequenced to a mean duplex coverage of &amp;gt;10000×. All expected mutations in the plasmid control titrations were identified for each titration. Both reproducibility and specificity in these samples were 1.0. The cell line controls were sequenced in a matrix of DNA input concentration vs Variant Allele Frequency (VAF). The Limit of Detection for the assay was a function of DNA concentration, with 0.4ng, 0.8ng and 3.3ng DNA required to identify mutations accurately and reproducibly at 0.05, 0.02 and 0.01 VAF respectively. The study found that the mutations detected among the human donor samples strongly correlated cumulative VAF with age (P value = 2.56×10−7). Conclusion This study describes the validation of a method for identifying low frequency variants in sun exposed skin. We demonstrate that the assay is a robust and reliable tool for detecting TP53 and NOTCH1 somatic variants associated with UVR exposure in normal-appearing skin down to 0.01 VAF with &amp;gt;0.8 sensitivity, specificity, and reproducibility. Damage from UV exposure is cumulative and therefore it is expected that older individuals will be more likely to have UVR-induced mutations in their skin, as we observed in this analysis. The assay can detect somatic DNA variations in normal-appearing skin for markers that are associated with UV damage and increased overall risk for certain nonmelanoma skin cancers.

Pemfigus Vulgaris

Pemphigus vulgaris (PV) is a chronic intraepidermal bullous disease that causes extensive skin lesions. Pemphigus vulgaris initially appears in the form of vesicles and loose-walled bullae that break easily on normal skin, which then develop into bullae with an erythematous base, then rupture, causing erosion, excoriation, which is usually accompanied by crusting and pain. Bullae occur due to damage or damage to intercellular adhesion due to IgG autoantibodies to desmoglein 3, causing epithelial cells called acantholysis (epidermal cell separation reaction). The hallmark of pemphigus is erosion that extends into normal skin by pulling on remnants of the blister wall or rubbing on the periphery of an active lesion. Erosion can be induced on normal-appearing skin away from the active lesion by mechanical stress or shear, a phenomenon known as the Nikolsky sign. Prognosis before corticosteroids are used, death occurs in 50% of patients within the first year. Treatment with corticosteroids makes the prognosis better. Pemphigus vulgaris in the early stages will be easier to control than those with widespread manifestations, the mortality rate will increase if there is a delay in therapy.

Whole transcriptome-based skin virome profiling in typical epidermodysplasia verruciformis reveals α-, β-, and γ-HPV infections.

HPVs are DNA viruses include approximately 450 types that are classified into 5 genera (α-, β-, γ-, μ-, and ν-HPV). The γ- and β-HPVs are present in low copy numbers in healthy individuals; however, in patients with an inborn error of immunity, certain species of β-HPVs can cause epidermodysplasia verruciformis (EV), manifesting as recalcitrant cutaneous warts and skin cancer. EV presents as either typical or atypical. Manifestations of typical EV are limited to the skin and are caused by abnormal keratinocyte-intrinsic immunity to β-HPVs due to pathogenic sequence variants in TMC6, TMC8, or CIB1. We applied a transcriptome-based computational pipeline, VirPy, to RNA extracted from normal-appearing skin and wart samples of patients with typical EV to explore the viral and human genetic determinants. In 26 patients, 9 distinct biallelic mutations were detected in TMC6, TMC8, and CIB1, 7 of which are previously unreported to our knowledge. Additionally, 20 different HPV species, including 3 α-HPVs, 16 β-HPVs, and 1 γ-HPV, were detected, 8 of which are reported here for the first time to our knowledge in patients with EV (β-HPV-37, -47, -80, -151, and -159; α-HPV-2 and -57; and γ-HPV-128). This study expands the TMC6, TMC8, and CIB1 sequence variant spectrum and implicates new HPV subtypes in the pathogenesis of typical EV.

Efficacy and safety of vixarelimab, a human monoclonal oncostatin M receptor β antibody, in moderate-to-severe prurigo nodularis: a randomised, double-blind, placebo-controlled, phase 2a study.

Prurigo nodularis is a chronic skin disease characterised by intensely pruritic, hyperkeratotic nodules. Vixarelimab, a human monoclonal antibody, binds to the beta subunit of the oncostatin M receptor, inhibiting signalling of both interleukin 31 and oncostatin M, two cytokine pathways that contribute to pruritus and nodule formation in prurigo nodularis. This double-blind, placebo-controlled, phase 2a trial was done at both private and academic dermatology outpatient research clinics across the United States and Canada (n=40). Patient eligibility criteria included age 18-75 years, physician-documented diagnosis of prurigo nodularis minimum 6 months duration of prurigo nodularis, presence of at least 10 pruritic nodules approximately 0.5-2cm in size on at least two different anatomical locations (excluding face and scalp) and involving the extremities, and presence of normal-appearing skin between nodules; atopic dermatitis as a comorbidity was exclusionary. Patients were required to have moderate-to-severe pruritus, defined as Worst Itch-Numeric Rating Scale (WI-NRS) score ≥7at screening and LS-mean weekly WI-NRS score ≥5 for each of the 2 consecutive weeks immediately before randomisation. Participants were randomly assigned (1:1) to receive weekly subcutaneous vixarelimab 360mg (720mg loading dose) or placebo using stratification factors (sex and presence of atopy) and block size 4 through the IWRS system. Stratification by atopy status was based on the reported high prevalence of atopy in this population and the potential impact of atopy in the immunopathologic process in prurigo nodularis. Patients, investigators, study sponsor, and site staff were masked to study treatment. The primary efficacy endpoint was least squares (LS)-mean percent change from baseline (PCFB) at Week 8 in weekly average Worst Itch-Numeric Rating Scale (WI-NRS) score. The primary efficacy endpoint was analysed with ANCOVA including treatment as fixed effect, with baseline WI-NRS, and randomisation stratification factor as covariates. All randomised patients who had at least 1 dose of study drug or placebo were included in the Safety Analysis Set. This trial is registered at ClinicalTrials.gov, NCT03816891. Of 50 patients randomised between March 11, 2019 and January 31, 2020, 23 vixarelimab recipients and 26 placebo recipients comprised the modified intent-to-treat analysis population (baseline LS-mean [SD] WI-NRS score, 8.3 [1.05]). Outcomes at Week 8 for vixarelimab versus placebo included LS-mean PCFB in WI-NRS score,-50.6% versus-29.4% (LS-mean difference [95% CI],-21.2% [-40.82,-1.60]; p=0.03); ≥4-point reduction in WI-NRS score, 52.2% (12/23) versus 30.8% (8/26) (p=0.11); PN-IGA score of 0 or 1, 30.4% (7/23) versus 7.7% (2/26) (p=0.03); LS-mean PCFB in pruritus VAS score,-54.4% versus-32.6% (p=0.03); and LS-mean PCFB sleep loss reduction (improvement),-56.3% versus-30.0% (p=0.02). No deaths, serious TEAEs, or TEAEs leading to dose interruption were reported. The percentage of vixarelimab recipients reporting any TEAE was 91.3% (21/23) versus 76.9% (20/26) of placebo recipients; drug-related TEAEs generally were similar between the two groups (vixarelimab, 43.5% [10/23]; placebo, 38.5% [10/26]). Vixarelimab demonstrated rapid reduction of pruritus and achievement of clear/almost clear skin in one-thirdof the patients by Week 8. Relief of itch and clearing of skin nodules represent two important potential therapeutic advances in the management of patients suffering from the debilitating disease Prurigo Nodularis. Kiniksa Pharmaceuticals, Ltd.

Multi-Omics Profiling of Skin Biopsies of Patients with Sclerodermatous Graft-Vs-Host Disease Suggests Therapeutic Potential of Targeting Don't Eat Me Signals

Introduction: Chronic graft-vs-host disease (cGVHD) is a major complication of allogeneic hematopoietic stem cell transplantation (HCT). This debilitating, chronic condition is characterized by inflammation, cell-mediated and humoral immunity, and ultimately tissue fibrosis. There is currently little or no understanding of the molecular pathogenesis of chronic cGVHD resulting in poor effective treatment strategies. Sclerodermatous GVHD (sclGVHD) is a highly morbid form of cGVHD associated with poor prognosis and low sensitivity to immune suppressive therapy. Understanding the underlying pathophysiology of sclGVHD and identifying the keyfibroticpathways involved iscriticalfor translation into targeted therapies that improve patient care. Methods: To further our understanding of the underlying pathophysiology of sclGVHD we used single cell RNA sequencing analyses on 5 fresh patient biopsy specimens and healthy controls. Human studies were carried out using: i) Patient skin biopsy samples (n =45) and matching them with 5 normal matched skin biopsy controls. These patient tissue samples were used to perform spatial gene expression analyses (10X Vizium) and identify the unique molecular profiles in each SclGVHD sample. ii) ATAC-sequencing analyses was performed on primary dermal fibroblasts from patients with SclGVHD to assess chromatin accessibility patterns genome wide in SclGVHD iii) TMA and tissues from patients were used to validate targets using immunofluorescent staining. iv) Animal studies were conducted to evaluate the therapeutic potential of an immune therapy-based treatment approach with a combined anti-fibrotic/anti-inflammatory strategy, and effectiveness of the treatment was determined with CyTOF studies. Results: To investigate cGVHD immunopathogenesis, normal-appearing skin, lesioned skin, and circulating immune cells from chronic patients were analyzed via single-cell RNA sequencing (figure A). A distinct transcriptional signature, the upregulation of clusters 11, 3, 1 and 17 in Scl cGVHD and the downregulation of clusters 6, 5, 9, 8 was observed. To validate these results in the context of tissue architecture we integrated spatial RNA sequencing on a tissue microarray assembled with 45 skin biopsies of SclGVHD patients to demonstrate a tight correlation between JUN activation and immune checkpoint upregulation (figure B).Hierarchical clustering of ATAC-seq signals revealed that the chromatin landscape was greatly remodeled after JUN deletion (figure C).We blocked CD47 and IL6 in the SclGVHD mouse model to evaluate therapeutic efficacy. We found a significant decrease of dermal thickness and collagen deposition in SclGVHD mice treated with blocking antibodies against IL6 and CD47 but not in untreated mice (figure D). We also detected a noticeable decrease in the expression of CD47, pJUN and IL6 in the treated group (figure E). Analyses of skin tissues from sclerotic lesions in mice using CyTOF revealed different cell populations in the skin clustered in spatially distinct groups. Finally, an increase in total leukocyte as well as a significant decrease in the fibroblast populations was observed (figure F). Conclusion: Our findings are significant because we profile skin biopsies matched with peripheral blood from patients with Scl GVHD with a multiomics approach and identify a unique molecular signature in patients with SclGVHD thus providing a rationale for subsequent combined interventions targeting innate immunity. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

4-Aminopyridine Induces Nerve Growth Factor to Improve Skin Wound Healing and Tissue Regeneration.

The discovery of ways to enhance skin wound healing is of great importance due to the frequency of skin lesions. We discovered that 4-aminopyridine (4-AP), a potassium channel blocker approved by the FDA for improving walking ability in multiple sclerosis, greatly enhances skin wound healing. Benefits included faster wound closure, restoration of normal-appearing skin architecture, and reinnervation. Hair follicle neogenesis within the healed wounds was increased, both histologically and by analysis of K15 and K17 expression. 4-AP increased levels of vimentin (fibroblasts) and alpha-smooth muscle actin (α-SMA, collagen-producing myofibroblasts) in the healed dermis. 4-AP also increased neuronal regeneration with increased numbers of axons and S100+ Schwann cells (SCs), and increased expression of SRY-Box Transcription Factor 10 (SOX10). Treatment also increased levels of transforming growth factor-β (TGF-β), substance P, and nerve growth factor (NGF), important promoters of wound healing. In vitro studies demonstrated that 4-AP induced nerve growth factor and enhanced proliferation and migration of human keratinocytes. Thus, 4-AP enhanced many of the key attributes of successful wound healing and offers a promising new approach to enhance skin wound healing and tissue regeneration.

Abstract 1909: Comparison of mutational burden hotspots in cutaneous squamous cell carcinoma and UV-exposed healthy skin for development of optimal targeted sequencing panels

Abstract Cutaneous squamous cell carcinoma (cSCC) is the second most common human malignancy in the United States. Chronic long-term ultraviolet (UV) exposure drives cSCC development. Early clonal mutation (CM) accumulation is the first step in photocarcinogenesis and is the first known manifestation of field cancerization. Understanding CM accumulation and development is expected to improve the prevention of field cancerization and cSCC. Nevertheless, CMs are poorly understood, and mutations in cSCC have yet to be systematically compared to CM in sun-exposed skin. Key steps for studies to detect and compare CMs are to design optimal targeted sequencing panels and compare mutational hotspot areas. The ideal panel should cover genomic regions containing maximum numbers of mutations within a given number of amplicons. Currently there is no computational tool to help optimize target area selection. Therefore, we aimed to create an algorithm to optimize sequencing target area design for capturing high mutation frequency hotspot areas. An R Shiny web application was created to identify an optimal sequencing target panel from an input mutation dataset and compare the distribution of mutational hotspots. The tool optimizes sequencing target areas based on preset amplicon length by identifying the best fitting panel of amplicons to capture mutations efficiently. Besides identifying optimal sequencing target areas, the developed software also efficiently identifies the most highly mutated genomic areas and compares target area overlaps. The developed algorithm was used to compare the mutational hotspots of cSCC and CM in clinically normal-appearing skin or cSCC from previous publications (cBioPortal, Martincorena et al, Hernando et al, Wei et al, Fowler et al). The current method was more efficient than tested previously available alternative methods by increasing target areas capture efficacy by 1.05 - 8.1-fold. Using the developed tool, we found that frequently mutated areas of CMs in normal skin significantly overlap (p &amp;lt; 0.005) with those of cSCC. Mutational hotspots of normal skin with a history of frequent UV exposure had 1.2-fold greater overlap with cSCC than skin with minimal UV exposure, suggesting the frequently UV-exposed skin carries a greater number of cSCC-related mutations. Although we found that normal skin CM hotspots better predict capture efficiency in sun-exposed normal skin than cSCC mutation hotspots, cSCC mutation hotspots can still design efficient sequencing targets in genomic areas where we do not have deep sequencing data on CM in normal skin. Our work provides a framework for helping design efficient, customized sequencing panels covering the genomic regions with the highest number of mutations. We expect the current algorithm to be a valuable tool for similar studies of other cancers. Citation Format: Sydney R. Grant, Megan E. Fitzgerald, Barbara A. Foster, Wendy J. Huss, Lei Wei, Gyorgy Paragh. Comparison of mutational burden hotspots in cutaneous squamous cell carcinoma and UV-exposed healthy skin for development of optimal targeted sequencing panels [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1909.

Combination of acitretin and narrowband UV-B for the treatment of vitiligo: A new treatment modality with an impact on tissue E-cadherin expression

Combination of acitretin and narrowband UV-B for the treatment of vitiligo: A new treatment modality with an impact on tissue E-cadherin expression

Nonlesional lupus skin contributes to inflammatory education of myeloid cells and primes for cutaneous inflammation.

Cutaneous lupus erythematosus (CLE) is a disfiguring and poorly understood condition frequently associated with systemic lupus. Previous studies suggest that nonlesional keratinocytes play a role in disease predisposition, but this has not been investigated in a comprehensive manner or in the context of other cell populations. To investigate CLE immunopathogenesis, normal-appearing skin, lesional skin, and circulating immune cells from lupus patients were analyzed via integrated single-cell RNA sequencing and spatial RNA sequencing. We demonstrate that normal-appearing skin of patients with lupus represents a type I interferon-rich, prelesional environment that skews gene transcription in all major skin cell types and markedly distorts predicted cell-cell communication networks. We also show that lupus-enriched CD16+ dendritic cells undergo robust interferon education in the skin, thereby gaining proinflammatory phenotypes. Together, our data provide a comprehensive characterization of lesional and nonlesional skin in lupus and suggest a role for skin education of CD16+ dendritic cells in CLE pathogenesis.

Disseminated tense bullae on newborn

Disseminated tense bullae on newborn

Evaluation of the healing activity of Cucurbita spp. leaf and seed extracts on experimental thermal burns

Burns are the most common skin injuries associated with many complications and high incidences of wound infections. Many compounds of plant products have been assessed to accelerate wound healing or reduce contamination. The possible healing effect of two Cucurbita spp. in skin burns was investigated. Cucurbita maxima and Cucurbita pepo leaves and seeds were extracted using ethanol and analyzed by gas chromatography–mass spectrometry. Second-degree burn wounds were made on laboratory rats, and the burn wounds were applied topically with extracts for 14 days and silver sulfadiazine (SS) for comparison. Serum vascular endothelial growth factor (VEGF) was analyzed post-treatment. Hydroxyproline content and histopathological changes were evaluated in skin biopsy. Phytochemical analysis displayed that C. maxima leaf extracts contain linolenic acid (21.94%), phytol (6.45%), and ascorbic acid (9.09%), and they were characterized by glucopyranoside (8.49%) and limonene (5.39%) compounds. The C. pepo leaves were characterized by palmitic acid (48.04%) and alkaloid (tetratriacontane: 13.11%). The seeds extracts of both Cucurbita spp. revealed a high ratio of linoleic acid, palmitic acid, and ascorbic acid. The leaf extracts accelerated wound contraction by 100%, with normal skin appearance, due to their more structural tissue layers, regular collagen fibers, ability to reduce inflammation, and high hydroxyproline content compared to the seed extracts and SS group. The bioactivity of unsaturated fatty acids, ascorbic acid, monoterpene, glycosides, and alkaloids provided the therapeutic property of these extracts. Serum VEGF was significantly reduced in the groups treated with extracts compared with those treated by the SS group. The leaf extracts of both Cucurbita species exhibited a healing efficiency on burn wounds, which were more effective than the seed extracts. Thus, they can be used as a promising plant derivative in burn treatments.

Frontal swelling – An unusual presentation of sarcoidosis: A case report

Subcutaneous sarcoidosis is a rare, cutaneous expression of systemic sarcoidosis. It is observed as asymptomatic firm, nodules covered by normal-appearing skin, principally on the extremities, which show the typical histopathology appearance of non-caseating granulomas localized to the subcutaneous tissue. It may be associated with the early benign, hilar lymphadenopathy of sarcoidosis or with the later stages of progressive sarcoidosis. Here, we report a case of subcutaneous sarcoidosis, which presented as an asymptomatic, forehead swelling with involvement of skull bone and lung. We report this case since subcutaneous sarcoidosis on the face with involvement of skull bone is a rarity.

Skin Erosions in a Newborn.

Skin Erosions in a Newborn.

214 Increased cutaneous activation of innate immunity and pro-apoptotic CXCR3B in patients with segmental vitiligo

Segmental vitiligo (SV) is a rare form of vitiligo, characterized by white patchy skin due to destruction of melanocytes affecting only one side of the body. The mechanisms are poorly understood. Recently we have demonstrated the importance of innate immunity and a newly discovered melanocytic chemokine receptor CXCR3B in triggering early anti-melanocytic response in non-segmental or generalized vitiligo. The role of innate immunity or CXCR3B in SV has not been studied. To do this, we obtained 2x4-mm skin biopsies from normal-appearing skin of SV patients (n=5), one from a non-depigmented skin of the affected side of the body (L) and the second from the contralateral non-effected side of the same patient (NL) and results were compared to healthy controls matched from age and biopsy location (n=5). Biopsies were cut in 2 for PCR and immunofluorescent staining. Our results have shown increased expression of vitiligo signature cytokines (IFNg) and chemokines (CXCL9 and CXCL10) as well as markers of ECM disruption (increased MMP-9 and decreased E-cadherin) in both L and NL sites. The targets of cellular stress (CXCL16, iHSP70) and cellular damage (mito-DNA) were only increased in L but not in NL sites. We found a significant increase in CXCR3B mRNA at both sites and a significantly increased number of CXCR3B+ melanocytes at L sites only, which may explain their increased responses to chemokines. In conclusion, our results suggest that SV patients, as has been described for NSV, have increased innate immunity throughout their skin which precedes depigmentation. Increased CXCR3B expression in SV suggests strategies fighting against the local stress or specifically targeting CXCR3B activation could provide effective approaches for halting the progression of this disabling disease.

Vascular injury in macroscopically normal skin of patients with severe COVID-19 infection: clinical-pathologic correlations

Abstract Objectives: Taking into account that the documentation of the histopathological features in severe disease caused by SARS-CoV-2 has been scarce due to the avoidance of performing autopsies, the aim of the study was to detect the microscopic changes associated with severe COVID-19 infection in normal-appearing skin, without prominent dermatologic signs of a generalized microvascular thrombotic disorder, in accordance with the clinical evolution of disease. Methods: In this morphological and immunohistochemical study we included cutaneous biopsy samples from 12 symptomatic patients with severe and critical type SARS-CoV-2 infection (with the admission date between February and June 2020), treated in the Intensive Therapy Unit Care of Emergency County Hospital Targu-Mures, Romania. Results: The average age of our patients was 65.18 ± 14.21 years (range 41 to 83), and 66.67% of the patients were male. The histological and immunohistochemical assessment of cutaneous biopsies: in 4 cases the histological examination revealed small fibrin thrombi in deep-seated venules and small veins of subcuticular adipose tissue, and also 4 cutaneous biopsies showed occlusive vascular thrombosis in association with massive perivascular inflammatory infiltrate destroying and compromising the integrity of the vessel wall. The immunohistochemical examination of the composition of perivascular inflammatory infiltrate showed a predominance of CD3 positive lymphocytes, admixed with CD68 positive Mo/ MF, some of them activated with FXIII expression. In the perivascular infiltrate, the presence of granulocytes and B lymphocytes was not characteristic. Conclusion: According to our observations, in severe COVID-19, the cutaneous tissue is involved even in the absence of clinically obvious changes. Due to the relatively easy accessibility of skin samples, these could be applied to determine the severity of the patient’s clinical status, and to predict the necessity for anti-complement or anticoagulant treatments in the early stages of a severe SARS-CoV-2 infection.

Increased Activation of Innate Immunity and Pro-Apoptotic CXCR3B in Normal-Appearing Skin on the Lesional Site of Patients with Segmental Vitiligo

The mechanisms involved in the pathogenesis of segmental vitiligo (SV) are poorly understood. Owing to its characteristic asymmetric unilateral distribution of depigmentation, proposed hypotheses include melanocytes and/or keratinocytes carrying mosaic mutation and being intrinsically abnormal and/or more susceptible to stress. Moreover, a subclinical inflammatory response is now well-demonstrated also in segmental forms of vitiligo (Speeckaert et al., 2020) and the involvement of CD8+ melanocyte-specific T-cell‒mediated immunity as is observed in generalized vitiligo (van Geel et al., 2010).